Cellular immunity and particularly CD4 T-cells have a central role in the control of Mycobacterium tuberculosis (Mtb) infection IFN-γ and TNF-α are thought to be crucial for protection against Mtb. Diagnosis of Mtb infection remains complex and requires several clinical, radiological, histo-pathological, bacteriological and molecular parameters. IFN-γ-release assays (IGRAs), i.e. Quantiferon and ELISpot, measure responses to antigens (e.g., ESAT-6 or CFP-10) that are mainly limited to Mtb, and discriminate infection from immunity induced by vaccination with Bacille Calmette-Guérin (BCG). IGRAs however do not discriminate between active disease and latent infection. While IFN-γ production alone showed no correlation with disease activity in chronic virus infection, polyfunctional (IFN-γ+IL-2+TNF-α) profiles of pathogen-specific T-cell responses have been correlated with disease activity. A definite correlation between active and latent Mtb infection, suitable for incorporation into an assay for differentiating between the two conditions, has not yet been described. Previous work has described a rough correlation between active disease and the presence of >50% of Mtb-reactive CD4 T cells producing TNF-α that do not also produce IFN-γ and IL-2 (e.g., TNF-α monospecific cells). However, this rough correlation was not sufficiently accurate or specific to serve as a true diagnostic tool. As described below, a specific correlation has been identified and an accurate, reproducible assay system for differentiating between active and latent infection by Mtb provided.